PROJECT 3: Neurobiological basis of negative-reinforcement drinking in female and male mice

项目3的作用是识别和解释“黑暗面”背后的神经生物学机制 在项目1和项目2中进行了研究。女性有更高的压力反应和更高的 抑郁症状比男性更严重，这可能是他们长期饮酒的可能性增加的原因。因为妇女 对压力对负强化饮酒的影响特别敏感（NRD，见整体部分）， 项目3的主要目的是确定与NRD和相关治疗相关的机制。杏仁核是一个 应激反应所必需的性二态结构，以及去甲肾上腺素（NE）和GABA信号传导 对压力诱发的行为至关重要基于GABA和NE在对两者的反应中的既定作用， 压力和酒精使用，我们假设，增加易感性慢性酒精使用和复发， 女性的性别差异部分是由于杏仁核次区域GABA-NE平衡的性别差异。我们已经表明 α2A受体的NE激动剂胍法辛降低杏仁核神经元的活性， 抗焦虑和抗抑郁样作用，具有神经元激活的性别依赖性模式。因此我们 假设靶向突触前和突触后NE受体或激活GABA神经元将抵消 NRD对应激引起的杏仁核神经元活动和行为有协同作用。我们 进一步假设这些神经机制与神经炎症通路相互作用，例如 小胶质细胞激活，以修改大脑区域的突触结构， 可能以性别依赖的方式改变NRD。最后，我们知道男性的酒精代谢是不同的， 女性，并且已经确定了与女性相比，女性大脑中乙醇代谢降低的影响更大。 雄性老鼠在项目3中，我们将1）确定靶向去甲肾上腺素能受体是否总体减少 乙醇摄入量，以及NRD在女性和男性小鼠，2）确定是否NE操作和GABA 杏仁核中的神经元活动对雌性和雄性小鼠的NRD具有协同作用， 遗传学以靶向特定的GABA能回路机制，3）确定去甲肾上腺素能受体的作用 和GABA神经元活性对乙醇诱导的小胶质细胞改变和突触密度的影响 小鼠，和4）确定脑酒精代谢和NRD中这些信号通路之间的相互作用。 这些研究将为项目1和2的研究提供相关的机制数据，以确定大脑 可能以性别依赖的方式调节与压力相关的酒精使用的机制，并确定如何 这导致了与酒精有关的行为的性别差异。

The role of Project 3 is to identify and translate the neurobiological mechanisms underlying the ‘dark side of addiction’ studied across Projects 1 and 2. Women have higher stress reactivity and higher rates of depressive symptoms than men that may underlie their increased likelihood of chronic drinking. Since women are particularly sensitive to effects of stress on negative reinforcement drinking (NRD, see Overall Section), a primary aim of Project 3 is to identify mechanisms related to NRD and related treatments. The amygdala is a sexually-dimorphic structure essential for stress reactivity, and both norepinephrine (NE) and GABA signaling are critical for stress-induced behaviors. Based on the established role of GABA and NE in response to both stress and alcohol use, we hypothesize that the increased susceptibility to chronic alcohol use and relapse in women is partly due to sex differences in GABA-NE balance in subregions of the amygdala. We have shown that guanfacine, a NE agonist at α2A receptors, decreases activity of amygdala neurons and induces anxiolytic and antidepressant-like effects, with sex-dependent patterns of neuronal activation. We therefore hypothesize that targeting pre- and postsynaptic NE receptors or activating GABA neurons will counteract NRD, and could have synergistic effects on amygdala neuronal activity and behaviors induced by stress. We further hypothesize that these neuronal mechanisms interact with neuroinflammatory pathways, such as microglial activation, to modify synaptic structure in the brain area, and that targeting these neuroadaptations could alter NRD in a sex-dependent manner. Finally, we know that alcohol metabolism differs in men and women, and have identified a greater effect of decreased brain metabolism of ethanol in female compared to male mice. In Project 3, we will 1) determine whether targeting noradrenergic receptors decrease overall ethanol intake, as well as NRD in female and male mice, 2) determine whether NE manipulations and GABA neuron activity in the amygdala have synergistic effects on NRD in female and male mice using molecular genetics to target specific GABAergic circuit mechanisms, 3) determine the effects of noradrenergic receptors and GABA neuron activity on ethanol-induced microglia alteration and synaptic density in female and male mice, and 4) identify interactions between brain alcohol metabolism and these signaling pathways in NRD. These studies will provide mechanistic data relevant for studies in Projects 1 and 2 to identify brain mechanisms that may modulate stress-related alcohol use in a sex-dependent manner, and to determine how this contributes to sex differences in alcohol-related behaviors.